The SMAC mimetic BV6 sensitizes colorectal cancer cells to ionizing radiation by interfering with DNA repair processes and enhancing apoptosis

• Published in: Radiation oncology (London, England) Vol. 10; no. 1; p. 198
• Main Authors: Hehlgans, Stephanie, Oppermann, Julius, Reichert, Sebastian, Fulda, Simone, Rödel, Claus, Rödel, Franz
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 17.09.2015; BioMed Central
• Subjects: Analysis; Apoptosis; Apoptosis - drug effects; Blotting, Western; Cancer; Cancer cells; Care and treatment; Cell Line, Tumor; Colorectal cancer; Colorectal Neoplasms - pathology; DNA; DNA damage; DNA repair; DNA Repair - drug effects; Flow Cytometry; Fluorescent Antibody Technique; Gene Knockdown Techniques; Genetic aspects; Humans; Inhibitor of Apoptosis Proteins - metabolism; Ionizing radiation; Medical equipment and supplies industry; Medical test kit industry; Oligopeptides - pharmacology; Patient outcomes; Proteins; Radiation-Sensitizing Agents - pharmacology; RNA; RNA, Small Interfering; Transfection; United States; Germany
• ISSN: 1748-717X; 1748-717X
• DOI: 10.1186/s13014-015-0507-4

In the present study, we aimed to investigate the effect of counteracting inhibitor of apoptosis (IAP) proteins using the small molecule Second Mitochondria-derived Activator of Caspase (SMAC) mimetic BV6 in combination with ionizing radiation on apoptosis, cell cycle regulation, DNA double-strand break (DSB) repair, three-dimensional (3D) clonogenic survival and expression of IAPs in colorectal carcinoma cells. Colorectal cancer cell lines (HCT-15, HT-29, SW480) were subjected to BV6 treatment (0-4 μM) with or without irradiation (2-8 Gy, single dose) followed by MTT, Caspase 3/7 activity, γH2AX/53BP1 foci assays, AnnexinV staining, cell cycle analysis, 3D colony forming assays and Western blotting (cellular IAP1 (cIAP1) and cIAP2, Survivin, X-linked IAP (XIAP)). BV6 treatment decreased cell viability and significantly increased irradiation-induced apoptosis as analyzed by Caspase 3/7 activity, AnnexinV-positive and subG1 phase cells. While basal 3D clonogenic survival was decreased in a cell line-dependent manner, BV6 significantly enhanced cellular radiosensitivity of all cell lines in a concentration-dependent manner and increased the number of radiation-induced γH2AX/53BP1-positive foci. Western blot analysis revealed a markedly reduced cIAP1 expression at 4 h after BV6 treatment in all cell lines, a substantial reduction of XIAP expression in SW480 and HT-29 cells at 24 h and a slightly decreased cIAP2 expression in HCT-15 cells at 48 h after treatment. Moreover, single or double knockdown of cIAP1 and XIAP resulted in significantly increased residual γH2AX/53BP1-positive foci 24 h after 2 Gy and radiosensitization relative to control small interfering RNA (siRNA)-treated cells. The SMAC mimetic BV6 induced apoptosis and hampered DNA damage repair to radiosensitize 3D grown colorectal cancer cells. Our results demonstrate IAP targeting as a promising strategy to counteract radiation resistance of colorectal cancer cells.